biostudies-literatureYang XDivision of ChemistryArmy Research OfficeNational Institute of General Medical SciencesNIGMS NIH HHSArmy Research Laboratory935-942https://www.ebi.ac.uk/biostudies/studies/S-EPMC8955669biostudies-literatureUnknown12(2)Vitamin B₁₂ derivatives catalyze a wide range of organic transformations, but B₁₂-dependent enzymes are underutilized in biocatalysis relative to other metalloenzymes. In this study, we engineered a variant of the transcription factor CarH, called CarH*, that catalyzes styrene C-H alkylation with improved yields (2-6.5-fold) and selectivity relative to cobalamin. While the native function of CarH involves transcription regulation via adenosylcobalamin (AdoCbl) Co(III)-carbon bond cleavage and <i>β</i>-hydride elimination to generate 4',5'-didehydroadenosine, CarH*-catalyzed styrene alkylation proceeds via non-native oxidative addition and olefin addition coupled with a native-like <i>β</i>-hydride elimination. Mechanistic studies on this reaction echo findings from eaACS catalysisControlling Non-Native Cobalamin Reactivity and Catalysis in the Transcription Factor CarH.PMC8955669R01 GM115665W911NF-19-1-0074CHE-1808133T32 GM131994Yang XJiang YZubi YSBaker LAGerroll BHRLewis JCKumar AfalseControlling Non-Native Cobalamin Reactivity and Catalysis in the Transcription Factor CarH.Vitamin B₁₂ derivatives catalyze a wide range of organic transformations, but B₁₂-dependent enzymes are underutilized in biocatalysis relative to other metalloenzymes. In this study, we engineered a variant of the transcription factor CarH, called CarH*, that catalyzes styrene C-H alkylation with improved yields (2-6.5-fold) and selectivity relative to cobalamin. While the native function of CarH involves transcription regulation via adenosylcobalamin (AdoCbl) Co(III)-carbon bond cleavage and <i>β</i>-hydride elimination to generate 4',5'-didehydroadenosine, CarH*-catalyzed styrene alkylation proceeds via non-native oxidative addition and olefin addition coupled with a native-like <i>β</i>-hydride elimination. Mechanistic studies on this reaction echo findings from ea2022-01-01T00:00:00Z2022 Jan2026-05-30T14:44:17.337Z2024-11-09T22:30:01.276ZS-EPMC89556693534076010.1021/acscatal.1c04748